(A) Experimental outline for comparing vaccine-elicited responses and immune protection. First, mice were made Ad5 seropositive by injecting them with 1 × 10⁹ PFUs of Ad5-Empty per mouse, once every 3 weeks for a total of 3 doses. These mice were then immunized intramuscularly with each respective vaccine based on the OVA antigen. After 4 weeks, mice were boosted homologously. (B) Summary of OVA-specific (K^b SIINFEKL⁺) CD8⁺ T cells in PBMCs. (C) Representative FACS plots showing OVA-specific CD8⁺ T cell in PBMCs. (D–G) Summary of OVA-specific CD8⁺ T cells in spleen (D), bone marrow (E), draining lymph nodes (F), and lungs (G). (H) OVA-specific antibody titers in sera. Data from C–H are from week ~8. (I) Experimental outline for comparing protective efficacy. (J and K) On day 3 after LM-OVA challenge, livers were harvested for H&E-stains (J) and measuring bacterial loads (K). Scale bars: 300 μm. Black arrows in J denote areas of necrosis. Data from 2 experiments (n = 4–5 mice per group/experiment). The vertical dashed line indicates the time of boosting. For B and D–H, indicated P values were determined by ordinary 1-way ANOVA test with Dunnett’s multiple comparisons (P values for B and H are from week 8). For K, indicated P values were determined by Kruskal-Wallis test with Dunn’s multiple-comparison test. Data are shown as mean ± SEM.